Plug and Play Anisotropy-Based Nanothermometers

Temperature is a crucial parameter in biology, nano-electronics, nanophotonics, and microfluidics. Optical methods excel for measuring temperature because they are noninvasive, spatially accurate, and can measure real time local changes in temperature. Among these, fluorescence anisotropy-based methods are particularly advantageous because they are less affected by changes in the probe concentration and irradiation conditions. However, at physiologically relevant temperature ranges in aqueous solution, fluorescence anisotropy contrast can only be achieved with rather large fluorescent proteins such as the green fluorescent protein (GFP), which can limit the range of applications through this method. Here, we propose a method to add thermosensitivity to any protein thereby transforming them into fluorescence anisotropy-based thermoprobes. It consists of covalently attaching a dye to the protein, which increases the rotational time of the dye-protein system compared to the free dye and confers thermosensitivity to the resulting bioconjugates. With this method we transformed bovine serum albumin, glucose oxidase and catalase into nanothermothers. This also allowed us to analyze the anisotropy signal changes occurring during the catalytic cycle of catalase, as well as their correlation with the reaction exothermicity. The potential of this method ensures applicability in extending temperature measurements to any protein-based experiments.